Vol 9, No 1 (2017)

Chimeric antigen receptors (CARs) are recombinant protein molecules that redirect cytotoxic lymphocytes toward malignant and other target cells. The high feasibility of manufacturing CAR-modified lymphocytes for the therapy of cancer has spurred the development and optimization of new CAR T cells directed against a broad range of target antigens. In this review, we describe the main structural and functional elements constituting a CAR, discuss the roles of these elements in modulating the anti-tumor activity of CAR T cells, and highlight alternative approaches to CAR engineering.

In our previous review, we described brain cholesterol metabolism in control conditions and in the case of some rare neurological pathologies linked to defects in the genes which are directly involved in the synthesis and/or traffic of cholesterol. Here, we have analyzed disruptions in cholesterol homeostasis in widespread neurodegenerative diseases (Alzheimer’s and Parkinson’s diseases) and autism spectrum disorders. We particularly focused on the synaptic dysfunctions that could arise from changes in both membrane cholesterol availability and oxysterol production. Notably, alterations in the brain cholesterol metabolism and neurotransmission occur in the early stages of these pathologies and the polymorphism of the genes associated with cholesterol homeostasis and synaptic communication affects the risk of onset and severity of these diseases. In addition, pharmacological and genetic manipulations of brain cholesterol homeostasis in animal models frequently have marked effects on the progression of neurodegenerative diseases. Thus, the development of Alzheimer’s, Parkinson’s and autism spectrum disorders may be partially associated with an imbalance of cholesterol homeostasis that leads to changes in the membrane cholesterol and oxysterol levels that, in turn, modulates key steps in the synaptic transmission.

Tumor response to therapeutic treatment is largely determined by its heterogeneity and the presence of intercellular junctions, hindering the penetration of large molecules deep into the three-dimensional structure of the tumor. In that context, 3D in vitro tumor models such as cancer cell spheroids are becoming increasingly popular. We obtained spheroids of human breast adenocarcinoma SKBR-3 overexpressing the HER2 cancer marker. The toxicity of HER2-targeted immunotoxin 4D5scFv-PE40 against spheroids was shown to be several orders of magnitude lower compared to a monolayer cell culture. The significant difference in the severity of the immunotoxin effect can be explained by the fact that it ineffectively penetrates the spheroid and predominantly influences the cells of the outer layers. The resulting tumor spheroid model can be used in development of drugs for targeted therapy as well as to study ways to improve the efficiency of anticancer agents by targeting cell-cell contacts.

This study was aimed at investigating the effect of prolyl-glycyl-proline (PGP) tripeptide on vascular permeability in rats with an inflammation. It was found that the peptide reduces the rat paw edema induced by a subcutaneous administration of histamine to the same extent as the conventional anti-inflammatory agent diclofenac. However, an assessment of the relative expression level of the cox-2 gene at the inflammation focus using real-time PCR showed that, in contrast to diclofenac, PGP does not affect the cox-2 gene expression. This is indicative of the fact that they have different mechanisms of action. We used the model of acute peritonitis induced by an intraperitoneal injection of thioglycolate to demonstrate that the inflammatory response of an organism is accompanied by increased vascular permeability in the tissues of the stomach and small intestine. Pre-administration (30 minutes before the induction of the inflammation) of PGP prevented this increase, whereby the level of vascular permeability, exudate volume in the peritoneal cavity, and the amount of the Evans Blue dye in this exudate remained at the control level. Therefore, these results suggest that the anti-inflammatory action of PGP is based on its ability to prevent an increase in vascular permeability.

Induced pluripotent stem cells (iPSCs) have the capacity to unlimitedly proliferate and differentiate into all types of somatic cells. This capacity makes them a valuable source of cells for research and clinical use. However, the type of cells to be reprogrammed, the selection of clones, and the various genetic manipulations during reprogramming may have an impact both on the properties of iPSCs and their differentiated derivatives. To assess this influence, we used isogenic lines of iPSCs obtained by reprogramming of three types of somatic cells differentiated from human embryonic stem cells. We showed that technical manipulations in vitro, such as cell sorting and selection of clones, did not lead to the bottleneck effect, and that isogenic iPSCs derived from different types of somatic cells did not differ in their ability to differentiate into the hematopoietic and neural directions. Thus, the type of somatic cells used for the generation of fully reprogrammed iPSCs is not important for the practical and scientific application of iPSCs.

Lytic transglycosylases are abundant peptidoglycan lysing enzymes that degrade the heteropolymers of bacterial cell walls in metabolic processes or in the course of a bacteriophage infection. The conventional catalytic mechanism of transglycosylases involves only the Glu or Asp residue. Endolysin gp144 of Pseudomonas aeruginosa bacteriophage phiKZ belongs to the family of Gram-negative transglycosylases with a modular composition and C-terminal location of the catalytic domain. Glu115 of gp144 performs the predicted role of a catalytic residue. However, replacement of this residue does not completely eliminate the activity of the mutant protein. Site-directed mutagenesis has revealed the participation of Tyr197 in the catalytic mechanism, as well as the presence of a second active site involving Glu178 and Tyr147. The existence of the dual active site was supported by computer modeling and monitoring of the molecular dynamics of the changes in the conformation and surface charge distribution as a consequence of point mutations.

The structure of the new enhancer En1A of the 1A region of the X chromosome of D. melanogaster was investigated. Two distinct regulatory elements were found. The first element is responsible for transcription activation, and the second element provides specific interaction with the promoter of the yellow gene. The findings support the hypothesis of a modular structure for enhancers, including certain sequences that bind transcription activators and special communication elements providing long-distance enhancer-promoter interaction.